美宁乐茶对小鼠睡眠调节作用的实验研究
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摘要
目的研究美宁乐茶对小鼠睡眠的调节作用及机制。方法将14只SPF级雄性C57BL/6J小鼠随机分为空白组和美宁乐茶组,各7只。小鼠每天分别自由饮用单蒸水和美宁乐茶浸出液(27 mg/mL)。1月后,采用Clock Lab监测系统观察小鼠昼夜活动,分析昼夜振幅、自由运动周期、相位偏移和每日活动度;采用RT-q PCR检测小鼠下丘脑视交叉上核(SCN)神经元和肝脏组织中生物时钟基因的表达;采用酶标仪法检测血清中谷丙转氨酶(ALT)、谷草转氨酶(AST)的活性及三酰甘油(TG)、总胆固醇(TC)、低密度脂蛋白胆固醇(LDL-C)、高密度脂蛋白胆固醇(HDL-C)的含量;采用血糖仪检测血糖水平。结果与空白组比较,美宁乐茶对小鼠的体质量,摄食量,血糖水平,血清ALT,AST活性及TG,LDL-C含量无显著影响,2组比较差异均无统计学意义(P>0.05),但可降低小鼠血清TC含量、LDL-C/HDL-C比值(P<0.05),并可显著降低小鼠每日活动度和昼夜振幅(P<0.01),延长自由运动周期及相位偏移(P<0.05或P<0.01),上调肝脏组织中Bmal1(brain and muscle ARNT-like 1)mRNA的表达(P<0.01),下调Dbp(albumin D site-binding protein)mRNA的表达(P<0.01),但对下丘脑SCN神经元时钟基因的表达无明显影响(P>0.05)。结论美宁乐茶可显著延长小鼠睡眠时间并提高睡眠质量,其作用可能是通过调控外周生物时钟基因的振荡性表达而实现的。
Objective To explore regulating effects of Meiningle Tea on the sleeping of mice and investigate the possible mechanisms. Methods 14 male C57 BL/6 J mice were randomly divided into control group and Meiningle Tea group,7 in each. Mice had free access to distilled water and Meiningle Tea water extraction( 27 mg/mL).After one month,the Clock Lab system was used to monitor diel activity of mice,as well as the amplitude,free-running period and phase shift of mouse circadian rhythms. RT-q PCR analysis was performed to determine clock gene expression levels in the hypothalamus suprachiasmatic nucleus( SCN) and liver. We also used ELISA to detect serum ALT and AST activity and levels of TG,TC,LDL-C and HDL-C. Finally,glucometer was employed to measure blood glucose concentrations. Results Compared with the control group,Meiningle Tea showed a modest effect on the body weight,food intake,blood glucose,serum ALT,AST activity,ATC levels,as well as LDL-C content of mice,there was no difference between two groups( P > 0. 05). But the tea significantly decreased serum TC levels and LDL-C/HDL-C ratio( P<0.05) and dramatically lowered locomotor activity and amplitude of mice( P<0.01),prolonged free-running period and reduced advanced shift of the clock phase( P<0.05 or P<0.01). At the molecular level,Meiningle Tea up-regulated hepatic Bmal1 mRNA expression levels( P < 0. 01),while down-regulated Dbp mRNA expression( P<0.01),but did not alter the expression of clock genes in the hypothalamus SCN neurons( P>0.05). Conclusion Meiningle Tea prolongs sleeping period and promotes sleeping quality through regulating circadian rhythms of systemic peripheral clock.
Objective To explore regulating effects of Meiningle Tea on the sleeping of mice and investigate the possible mechanisms. Methods 14 male C57 BL/6 J mice were randomly divided into control group and Meiningle Tea group,7 in each. Mice had free access to distilled water and Meiningle Tea water extraction( 27 mg/mL).After one month,the Clock Lab system was used to monitor diel activity of mice,as well as the amplitude,free-running period and phase shift of mouse circadian rhythms. RT-q PCR analysis was performed to determine clock gene expression levels in the hypothalamus suprachiasmatic nucleus( SCN) and liver. We also used ELISA to detect serum ALT and AST activity and levels of TG,TC,LDL-C and HDL-C. Finally,glucometer was employed to measure blood glucose concentrations. Results Compared with the control group,Meiningle Tea showed a modest effect on the body weight,food intake,blood glucose,serum ALT,AST activity,ATC levels,as well as LDL-C content of mice,there was no difference between two groups( P > 0. 05). But the tea significantly decreased serum TC levels and LDL-C/HDL-C ratio( P<0.05) and dramatically lowered locomotor activity and amplitude of mice( P<0.01),prolonged free-running period and reduced advanced shift of the clock phase( P<0.05 or P<0.01). At the molecular level,Meiningle Tea up-regulated hepatic Bmal1 mRNA expression levels( P < 0. 01),while down-regulated Dbp mRNA expression( P<0.01),but did not alter the expression of clock genes in the hypothalamus SCN neurons( P>0.05). Conclusion Meiningle Tea prolongs sleeping period and promotes sleeping quality through regulating circadian rhythms of systemic peripheral clock.
引文
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[12]Moller-Levet C S,Archer S N,Bucca G,et al.Effects of insufficient sleep on circadian rhythmicity and expression amplitude of the human blood transcriptome[J].Proc Natl Acad Sci USA,2013,110(12):1 132-1 141.
[2]Uth K,Sleigh R.Deregulation of the circadian clock constitutes a significant factor in tumorigenesis:a clockwork cancer[J].Biotechnology&Biotechnological Equipment,2014,28(3):379-386.
[3]麻砚涛,罗浑金,靳倩,等.O-Glc NAc修饰调节生物节律研究进展[J].生命科学,2015,27(11):1 403-1 408.
[4]张晓萍.失眠的自我调节与综合治疗[J].中国实用医药,2009,4(2):209-210.
[5]张莹.NHBA和B2镇静睡眠作用及其机制研究[D].北京:中国医学科学院药物研究所,2010.
[6]黄维,金邦荃,程光宇,等.酸枣仁功效成分测定及改善睡眠保健功能的研究[J].时珍国医国药,2008,19(5):1 173-1 175.
[7]黎同明,高洁,王桂香.乌梅水煎液镇静催眠及抗惊厥作用实验研究[J].中医学报,2011,26(7):818-820.
[8]杨淑芬,杨艺娇,王海颖.浅述开窍醒脑中药作用机制[J].中国中医急症,2017,26(2):244-260.
[9]安扬,徐璎.哺乳动物昼夜节律机制研究进展[J].生命科学,2015,27(11):1 372-1 379.
[10]周岚,曹济民.小鼠转轮运动昼夜节律研究技术的建立[J].医学研究杂志,2011,40(6):31-35.
[11]孙秀杰,陈亚琼,刘浥.进食行为调控外周生物钟节律的研究进展[J].生命科学,2015,27(11):1 386-1 391.
[12]Moller-Levet C S,Archer S N,Bucca G,et al.Effects of insufficient sleep on circadian rhythmicity and expression amplitude of the human blood transcriptome[J].Proc Natl Acad Sci USA,2013,110(12):1 132-1 141.